PROJECT 2 PROJECT SUMMARY/ABSTRACT Loss of the CFTR anion channel causes cystic fibrosis (CF), which is characterized by bacterial infection and inflammation. To understand how CF disrupts airway defenses, we generated CF pigs and found that on the day they are born, they have impaired respiratory host defenses. Within weeks, they spontaneously develop hallmarks of CF airways: infection, inflammation, remodeling, mucus accumulation, and obstruction. Studies of CF pigs revealed at least two respiratory host defense defects: reduced activity of antimicrobials in airway surface liquid and impaired mucociliary transport. Both defects are caused, at least in part, by an abnormally acidic airway liquid. Loss of CFTR-mediated bicarbonate secretion leaves unchecked acid secretion that reduces pH. We found that a proton pump (ATP12A) is responsible for secreting acid into the airways. The overarching goal of this project is to understand how ATP12A acid secretion impairs host defense and whether the acidic pH causes CF lung disease. The project addresses three main questions. First, how is ATP12A acid secretion regulated in CF airways? Understanding the control of ATP12A is important because varying the rate of H+ secretion will change pH, which could exacerbate or attenuate disease severity. Second what is the pH in the lumen of submucosal glands? Submucosal glands produce most of the airway mucus. Earlier work showed that mucus emerging from CF submucosal glands ducts sometimes failed to break free from the duct, which impaired mucociliary transport. Those discoveries and knowledge that an acidic pH increases mucus elasticity make it imperative to know the pH in the submucosal gland lumen. This proposal develops novel means to obtain that information. Third, does disrupting the ATP12A gene rescue CF defects? CF pigs that have a disrupted ATP12A gene will allow a test of the hypotheses that preventing acid secretion will increase pH in the submucosal gland lumen, improve mucociliary transport, and prevent CF lung disease. This research will allow us to better understand how CF alters pH in the submucosal gland lumen and at the airway surface, thereby disrupting respiratory host defenses. The knowledge of CF pathogenesis will accelerate discovery of novel therapies and cures for this lethal disease.